The long-term goal of this translational research project is to improve survival of HIV/HCV co-infected patients by finding new pharmaceutical targets in HCV and identifying new viral products that can be used in vaccines and in diagnostic and prognostic tests. HCV-associated liver disease is a leading cause of death among HIV/HCV co-infected patients. Using custom bioinformatics tools, we found several RNA structural elements, and of utmost significance, discovered an alternate reading frame, whose products, ARFPs, stimulate immune responses in both mono-infected and co-infected patients (Walewski et al., RNA, 2001 and 2002). A. Experiments in Specific Aim I test the hypothesis that ARF expression and the immune response to ARFPs are modulated during the course of disease progression and by treatment with interferon. ELISAs will be used to detect and titer anti-ARFP antibodies in co-infected and mono-infected patients. Proliferation assays will be used to measure T cell responses. Western blots will be used to determine the sizes of core/ARFP proteins in both non-tumor and HCC liver specimens; EIA and QRT-PCR will be used to quantify core antigen and HCV RNA levels, respectively. Immunohistochemistry will be used to localize core protein and ARFPs in liver specimens. B. Experiments in Specific Aim II test the hypothesis that ARF expression is regulated by RNA structures that stimulate recoding events. ARF is in the +1 reading frame. Expression of other +1 reading frame genes that encode auto-regulatory factors, such as antizyme, is stimulated by specific RNA structural elements, which are similar to those in the core/ARF gene region. We will use in vitro translation and directed mutagenesis to determine the role of RNA elements in ARF expression. C. Experiments in Specific Aim III test the hypothesis that core and ARFPs have distinct biological properties, and that ARFPs stimulate cell growth. Gene expression profiling, cell cycle measurements, and oxidative stress responses will be used to differentiate the effects of core and ARFPs in transfected Tet-Off HepG2 cells. Western blots, immunoprecipitation, and mass spectroscopy will be used to determine the sizes of core/ARF gene products. ARFP-specific poly- and monoclonal antibodies will be developed to determine ARFPs' subcellular location. The yeast two-hybrid system will identify human proteins that bind ARFPs. These studies will identify new targets for therapeutics and vaccine development.

National Institute of Health (NIH)
National Institute on Drug Abuse (NIDA)
Research Project (R01)
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Study Section
Special Emphasis Panel (ZRG1-AARR-4 (02))
Program Officer
Khalsa, Jagjitsingh H
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Mount Sinai School of Medicine
Internal Medicine/Medicine
Schools of Medicine
New York
United States
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Vachon, Marie-Louise C; Factor, Stephanie H; Branch, Andrea D et al. (2011) Insulin resistance predicts re-treatment failure in an efficacy study of peginterferon-?-2a and ribavirin in HIV/HCV co-infected patients. J Hepatol 54:41-7
Gutierrez, Julio A; Parikh, Neil; Branch, Andrea D (2011) Classical and emerging roles of vitamin D in hepatitis C virus infection. Semin Liver Dis 31:387-98
Gutierrez, Julio A; Klepper, Arielle L; Garber, John et al. (2011) Cross-genotypic polyclonal anti-HCV antibodies from human ascitic fluid. J Virol Methods 171:169-75
Branch, Andrea D; Rice, Charles M (2010) Antisense gets a grip on miR-122 in chimpanzees. Sci Transl Med 2:13ps1
Childs, Kathryn E; Fishman, Sarah L; Constable, Catherine et al. (2010) Short communication: Inadequate vitamin D exacerbates parathyroid hormone elevations in tenofovir users. AIDS Res Hum Retroviruses 26:855-9
Chang, Theresa L; Klepper, Arielle; Ding, Jian et al. (2010) Human peritoneal macrophages from ascitic fluid can be infected by a broad range of HIV-1 isolates. J Acquir Immune Defic Syndr 53:292-302
Eng, Francis J; Walewski, Jose L; Klepper, Arielle L et al. (2009) Internal initiation stimulates production of p8 minicore, a member of a newly discovered family of hepatitis C virus core protein isoforms. J Virol 83:3104-14
Cox, Andrea L; Page, Kimberly; Bruneau, Julie et al. (2009) Rare birds in North America: acute hepatitis C cohorts. Gastroenterology 136:26-31
Fishman, Sarah L; Factor, Stephanie H; Balestrieri, Cinzia et al. (2009) Mutations in the hepatitis C virus core gene are associated with advanced liver disease and hepatocellular carcinoma. Clin Cancer Res 15:3205-13
Fishman, Sarah L; Murray, Jacinta M; Eng, Francis J et al. (2008) Molecular and bioinformatic evidence of hepatitis C virus evolution in brain. J Infect Dis 197:597-607

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